Research conducted in the current project over the past year in the Neurophysiological Pharmacology Section has continued to focus on the sources and consequences of changes in basal ganglia function in Parkinsons disease (PD). As previously reported, we have developed a strategy for studying ongoing activity in motor circuits while the rat performs a task which allows us to quantitate parkinsonian motor deficits. We have accomplished this through the use of a circular treadmill. A paddle is lowered over the rotating circular track to encourage the rat to keep walking. A rat with a unilateral dopamine cell lesion - mimicking unilateral PD - can walk effectively on this treadmill at a slow speed as long as he is oriented in the direction ipsilateral to the dopamine cell lesion, with the paws opposite the lesioned hemisphere on the outside of the circular track. His ability to walk in the opposite contralateral direction with his affected paws on the inside of the track is typically very limited, and provides a read-out of the motor disability. The treadmill setup allows us to monitor basal ganglia circuit spiking and local field potential (LFP) activity continuously from the intact and lesioned hemispheres as the animal walks and rests. In the past year, we have focused on addressing four questions. 1.Is there evidence for a causal relationship between motor deficits and increased beta range synchronization in the basal ganglia circuits? Our data fails to show a linear relationship between increases in LFP power in the beta range in basal ganglia thalamocortical circuits and the emergence of bradykinesia. In the hemiparkinsonian rat model, rats were implanted with electrode bundles in various parts of the basal ganglia and a cannula was inserted into the area over the median forebrain bundle to allow continuous recordings shortly after infusion of the toxin which kills dopamine neurons: 6-hdorxydopamine (6-OHDA). A significant trend toward increasing LFP power in the high beta (30 35Hz) frequency range emerges over time post lesion when the rat was walking in the ipsiversive direction, leveling off between 1 and 2 weeks post lesion. However, this increase in high beta range oscillatory activity does not reach significance, relative to baseline, in the motor cortex and substantia nigra until about 4 days after 6-OHDS infusion of the toxin. Coherence between high beta range oscillatory activity in the motor cortex and substantia nigra becomes significant slightly earlier, at 3 days post lesion. In contrast, motor deficits are fully evident within hours after the 6-OHDA-mediated dopamine cell lesion in hemiparkinsonian rats, as the rat fails to walk effectively in the contraversive direction on the treadmill, and does poorly on stepping tests. A second strategy for studying the effect of loss of dopamine receptor stimulation, treatment twice daily with D1/D2 dopamine receptor antagonists, provided similar results. Catalepsy is evident very shortly after the dopamine antagonist treatment, and wears off 4 or 5 hours after each injection. High beta oscillatory LFP activity emerges in the motor cortex and SNpr only gradually after several days when the animal is moved passively, and does not wear off between drug treatments, as does the catalepsy. By the end of the week, high beta range activity in maintained throughout the periods between the twice daily injections if the animal is actively walking on a treadmill. Thus, the emergence of the high beta oscillatory activity is not well correlated with the expression of catalepsy. A third strategy for attempting to correlate catalepsy induced by loss of dopamine was the use of unilateral infusion of tetrodotoxin into the median forebrain bundle. Motor deficits emerged in these rats in the absence of increases in beta range activity. Overall, these observations indicate that either a very small, difficult to measure change in synchronized activity in basal ganglia circuits is sufficient to induce marked motor deficits, or, some other consequence of loss of dopamine receptor stimulation is more directly responsible for the rapid emergence of motor deficits associated with dopamine cell death. 2. What changes in spiking and local field signaling in basal ganglia thalamocortical circuits correlate most closely with the emergence of motor deficits induced by decreases in dopamine receptor stimulation? In contrast to the slow changes in exaggerated beta range activity which emerge over time in basal ganglia thalamocortical circuits after blockade of dopamine receptors, a rapid change was noted in the motor cortex in our studies. This early change did correlate more closely with the emergent bradykinesia. In all models studied, there was an early and intriguing shift in peak frequency of the LFP recorded in the motor cortex when hemiparkinsonian rats walked on the circular treadmill. Over the first hours post lesion, as the rats show difficulty walking, the peak LFP frequency in the motor cortex shifts 40 Hz toward the 30 Hz range. Administration of a dopamine agonist within these first hours reversed this decrease in motor cortical rhythm as do other manipulations which improve motor activity. Pilot studies are showing that infusion of drugs which either stimulate or block the receptors for the inhibitory transmitter GABA in the subthalamic nucleus can briefly restore treadmill walking in the hemiparkinsonian rat and reverse the change in cortical frequency. Current results suggest that the critical changes leading to motor deficits do involve changes occurring throughout the basal ganglia, but it remains to be determined how and where locomotor centers are impacted. 3. Can pharmacogenetic tools provide insight into critical nodes in basal ganglia thalamocortical circuits associated with motor dysfunction in Parkinsons disease? To probe for further insight into how the basal ganglia becomes dysfunctional after loss of dopamine and how, in a broader sense, brain circuits become aberrantly synchronized, we have begun studies involving infusion of the inhibitory DREADD (Designer Receptors Exclusively Activated by Designer Drugs) virus, AAV2/8-hSyn-Hm4d(Gi)-mCjerry, into the globus pallidus and substantia nigra to evaluate the utility of this approach in manipulating activity in different components of the motor circuit. In particular, data is emerging from studies in the globus pallidus and the substantia nigra suggesting the existence of multiple subtypes which may be differentiated by differences in protein expression and anatomical connections. Parvalbumin (PV)-iCre rats are now available and we are hoping to use them together with the DREADD viruses to study the effects of selectively inhibiting PV expressing interneurons in the brain motor circuits. These studies will help us understand how to better compensate for the dysfunctional changes that emerge in neuronal activity in Parkinsons disease. 4. Can we use our combination of neurological and motor function tests to characterize novel treatments for Parkinsons disease? We are currently testing a novel dopamine D3 receptor agonist in our Parkinsonian rats. Tests involve testing the ability of this new drug to reduce both neurophysiological and behavioral aspects of parkinsonian symptoms without inducing dyskinesia. It has previously been difficult to conduct such a study as drugs with affinity for D3 receptors have typically also been effective at D2 receptors. This study, conducted in collaboration with Dr. David Sibley, who has developed this drug, will show, for the first time, whether an agent highly selective for the D3 dopamine receptor subtype will reduce motor deficits without inducing dyskinesias, a common side effect of drugs used to treat Parkinsons disease.